The goal of this project is to discover and develop novel broad-spectrum antibiotics acting against critically important human pathogens. The unmet need is especially acute for Gram-negative pathogens, where we are rapidly running out of options for treating multidrug resistant species of Pseudomonas, Acinetobacter, Burkholderia, Klebsiella and Enterobacter. This is a challenging goal, since the last novel class of broad-spectrum antibiotics was discovered 40 years ago. Currently, there are no broad-spectrum leads in the global pipeline of drug discovery, according to the review of this important subject at the last ICAAC meeting. The main barrier to progress in antimicrobial drug discovery is the lack of sources for novel compounds. NovoBiotic has a proprietary technology capable of meeting this challenge. We are obtaining new antimicrobials from a previously inaccessible source - uncultured fungi. In Phase I, we developed a trap for specific capture and growth of uncultured fungi. Many antimicrobials, including ?-lactams and cephalosporins derive from culturable representatives of this group of microorganisms. The trap consists of two layers of sterile agar sandwiched between semi-permeable membranes that is inserted into soil. Fungi produce hyphae which enable them to penetrate into the trap through the pores in the membrane, into the upper layer of agar and form pure colonies. Many uncultured species that initially grow in situ are then capable of growing under standard laboratory conditions in vitro. The aims of Phase I were to assemble a collection of 200 fungi using the trap method, screen them for antimicrobial activity and dereplicate 25 of them. We achieved these milestones and discovered a novel broad- spectrum antimicrobial, Novo15, from an uncultured fungus only distantly related to known organisms. In Phase II we will validate the compounds discovered in Phase I in in vitro biological assays and animal models of S. aureus and K. pneumonia pulmonary infection. Also, we will isolate and screen an additional 250,000 fungal strains from the trap (Aim 1), validate potential leads using in vitro biological tests and determine the structure of promising compounds (Aim 2), and perform animal tests on 10 novel compounds (Aim 3). The end result of Phase II will be 2-3 validated, novel broad-spectrum antimicrobials. These will be developed into drugs in the next phase of the program. PUBLIC HEALTH RELEVANCE: There is increasing public health concern about the ability of currently available antibiotics to keep pace with evolving microbial resistance; naturally acquired in the case of community and hospital based infections, and potentially engineered in the case of bioterror agents. One solution to this problem is the development of new antibiotics. The purpose of this project is to use our new technology to identify new classes of antibiotics with broad-spectrum activity against both conventional and biodefense pathogens.